腾冲嗜酸热两面菌伴侣素ATcpnβ底物结合特性研究
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国家自然科学基金(30770496),国家高技术研究发展计划(863)(2007AA100604)和国家重点基础研究发展计划(973)(2006CB503910, 2006CB806506)资助项目


Substrate Binding Properties of Thermosome ATcpnβ From Acidianus Tengchongensis
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This work was supported by grants from The National Natural Science Foundation of China (30770496), Hi-Tech Research and Development Program of China (2007AA100604) and National Basic Research Program of China (2006CB503910, 2006CB806506)

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    摘要:

    嗜酸嗜热古菌腾冲嗜酸热两面菌(Acidianus tengchongensis)来源的Ⅱ型伴侣素ATcpnβ已获得晶体结构解析,其顶端结构域突触下端相应于Ⅰ型伴侣素GroEL的重要底物结合位点处的氨基酸多为极性氨基酸,将其突变为疏水性氨基酸时,突变体对变性底物的捕获能力显著增强.表面等离子共振研究表明,ATcpnβ对于化学变性底物的再折叠中间体的其捕获作用不依赖于Mg2+/ATP.前期对该伴侣素冷冻电镜观察和结构解析表明,ATP的存在并不能驱动ATcpnβ从开放构型向闭合构型转变,但是表面等离子共振研究表明,ATcpnβ对热变性过程中已经聚集的底物的捕获作用依赖于Mg2+/ATP,说明Mg2+/ATP可以介导ATcpnβ顶端结构域一定的构象变化,引起顶端结构域疏水残基的进一步暴露,从而能够与大分子聚集体紧密结合.两个方面的研究均表明,伴侣素蛋白与变性底物的结合仍然以疏水相互作用为主,并且伴侣素蛋白与变性底物的结合受Mg2+/ATP的结合调控,与伴侣素蛋白疏水面的暴露程度相关.

    Abstract:

    The residues associated with substrate binding in GroupⅠ chaperonin GroEL are hydrophobic. However, the corresponding residues in groupⅡ chaperonin ATcpnβ from Acidianus tengchongensis are hydrophilic.When these hydrophilic residues in ATcpnβ were mutated to hydrophobic residues, i.e. residues 236 from G to Y, 237 from M to F, 288 from D to L, 295 from A to V, 313 from A to V and 314 from K to V, they enhanced the inhibitory effect of ATcpnβ on the refolding of acid-denatured GFP and the inhibition activity of citrate synthase (CS) thermal aggregation. Hydrophobic interaction may contribute more to peptides binding affinity both in groupⅠ and groupⅡ chaperonin. Chaperonins have been proved to have ATP-dependent peptides refolding ability. However, it is still unclear whether peptides binding ability of chaperonins is ATP-dependent. Surface plasma resonance (SPR) analysis is used to test chaperonin binding ability to different peptides with different denaturing level. These assays revealed that ATcpnβ could capture guanidine hydrochloride denatured malate dehydrogenase in a Mg2+/ATP independent manner while it bind thermal aggregated citrate synthase or lysozyme in a Mg2+/ATP dependent manner. It has been proposed that chaperonin conformatinal changes induced by Mg2+/ATP to expose more hydrophobic surface is required for chaperonin capturing thermal aggregated peptides which has larger hydrophobic surfaces.

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王丽,张凯,樊峥,董志扬,孙飞.腾冲嗜酸热两面菌伴侣素ATcpnβ底物结合特性研究[J].生物化学与生物物理进展,2011,38(2):151-158

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  • 收稿日期:2010-09-28
  • 最后修改日期:2010-11-22
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  • 在线发布日期: 2010-11-23
  • 出版日期: 2011-02-20